The broad, long term objective of this research proposal is to understand the process of epididymal maturation of sperm. this poorly defined process is essential for fertilization; sperm that have entered the cauda epididymis and vas deferens are able, after a period of capacitation, to fertilize eggs whereas sperm taken from the testis or caput epididymis are not. The working model is that sperm maturation within the epididymis is not a single event but involves sequential, progressive in sperm properties brought about by the transduction of signals from the epididymal secretions to the intracellular spaces of the sperm. The challenge in understanding sperm maturation in the epididymis, therefore, is one of determining the modifications to sperm components that are essential for fertilization, characterizing how these components become functional as a result of epididymis-induced alterations, and elucidating the epididymal factors or conditions that stimulate these changes. This proposed research will examine changes that take place within the confines of the important sperm acrosome, the acrosome. The alterations to be investigated are the structural and functional modifications of the sperm-specific serine protease zymogen, proacrosin, and structural changes to another acrosomal glycoprotein, acrogranin. specific Aim 1 will characterize, using high performance liquid chromatography for carbohydrate analysis, the modification of proacrosin oligosaccharides that takes place in the corpus epididymis. Specific Aim 2 will determine how these alterations affect enzymatic activity. Specific Aim 3 will determine where specific cleavages in the polypeptide backbone of acrogranin occur by partial amino acid sequencing of products isolated from cauda epididymal sperm. Specific Aim 4 will use the biochemical information from Specific Aims 1 and 3 to predict and to identify the specific glycosidase(s) and protease(s) responsible for the epididymis-induced alterations of proacrosin and acrogranin. Specific Aim 5 is to develop a bioassay using the modifications of proacrosin and acrogranin as markers of epididymal maturation. this bioassay will be used to determine the conditions or substances that initiate these changes in testicular and caput epididymal sperm. These experiments will provide a unique approach to investigate epididymal maturation of sperm. The bioassay developed during the proposed project will be used in future studies to characterize the conditions or purify the factors responsible for initiating this aspect of sperm maturation. Furthermore, the information gained from these studies will find application in the treatment of male infertility caused by blockage of the epididymis or vas deferens. New approaches toward development of a male contraceptive may also result from the information gained from these studies.